Abstract
Protein kinase Czeta (PKCzeta) is an intracellular serine/threonine protein kinase that has been implicated in the signaling pathways for certain inflammatory cytokines, including interleukin-1 (IL-1) and tumor necrosis factor alpha (TNF-alpha), in some cell types. A study of gene expression in articular chondrocytes from osteoarthritis (OA) patients revealed that PKCzeta is transcriptionally up-regulated in human OA articular cartilage clinical samples. This finding led to the hypothesis that PKCzeta may be an important signaling component of cytokine-mediated cartilage matrix destruction in articular chondrocytes, believed to be an underlying factor in the pathophysiology of OA. IL-1 treatment of chondrocytes in culture resulted in rapidly increased phosphorylation of PKCzeta, implicating PKCzeta activation in the signaling pathway. Chondrocyte cell-based assays were used to evaluate the contribution of PKCzeta activity in NF-kappaB activation and extracellular matrix degradation mediated by IL-1, TNF, or sphingomyelinase. In primary chondrocytes, IL-1 and TNF-alpha caused an increase in NF-kappaB activity resulting in induction of aggrecanase-1 and aggrecanase-2 expression, with consequent increased proteoglycan degradation. This effect was blocked by the pan-specific PKC inhibitors RO 31-8220 and bisindolylmaleimide I, partially blocked by Gö 6976, and was unaffected by the PKCzeta-sparing inhibitor calphostin C. A cell-permeable PKCzeta pseudosubstrate peptide inhibitor was capable of blocking TNFand IL-1-mediated NF-kappaB activation and proteoglycan degradation in chondrocyte pellet cultures. In addition, overexpression of a dominant negative PKCzeta protein effectively prevented cytokine-mediated NF-kappaB activation in primary chondrocytes. These data implicate PKCzeta as a necessary component of the IL-1 and TNF signaling pathways in chondrocytes that result in catabolic destruction of extracellular matrix proteins in osteoarthritic cartilage.
Highlights
protein kinase C (PKC) Function in Chondrocytes intervention points that could mitigate the destruction of articular cartilage in OA
We find that nuclear factor B (NF-B) activation by tumor necrosis factor (TNF) and IL-1 in chondrocytes requires PKC activity and, most importantly, that inhibition of PKC blocks cytokinemediated up-regulation of aggrecanase expression and the resulting destruction of articular cartilage extracellular matrix proteoglycans that is a hallmark of the OA disease process
Protein Kinase C Is Transcriptionally Up-regulated in Human Osteoarthritic Cartilage—Gene expression changes in articular chondrocytes from lesional (n ϭ 14) and adjacent nonlesional (n ϭ 13) osteoarthritic cartilages compared with nonosteoarthritic cartilages (n ϭ 10) were analyzed using Affymetrix GeneChip U95Av.2 arrays
Summary
PKC Function in Chondrocytes intervention points that could mitigate the destruction of articular cartilage in OA. Following pretreatment with inhibitors (control culof PKC phosphorylation in the IL-1␣-treated primary tures received no inhibitor pretreatment), TNF-␣ or bovine chondrocytes were very similar to T/C-28a2 cells that IL-1␣(or no cytokine) was added to the cultures.
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